Lung cancer is a leading cause of cancer death in the world. Conventional cytotoxic-based chemotherapy regimens are at their limits of maximal effectiveness. New treatment paradigms incorporating inhibitors to specific driver mutations, such as EGFR and ALK, have shown improved clinical benefits. These new treatments depend on identification and evaluation of clinically relevant genetic lesions that drive and maintain the malignancy. Lung cancer is a heterogeneous disease. Clinically relevant mutations in ~50% of non-small cell lung cancer (NSCLC) remain to be identified and evaluated. It is envisioned that recent advances in genomic technologies will accelerate the discovery of new genetic alternations in human cancer. An important aspect in translating the new genomic findings into improved treatments is to have in vivo preclinical models that faithfully phenocopy the newly identified genes in the human diseases for evaluating drug response and resistance. Recurrent gene fusion between kinesin family member 5B (KIF5B) and the RET tyrosine kinase (KIF5B-RET) is the most recent genetic alternation identified in human lung adenocarcinoma. Preclinical models of KIF5B-RET have been limited to xenografts of transfected NIH3T3 fibroblasts and Ba/F3 pre-B- lymphocytes. The goal of this study is to generate doxycycline-inducible, lung-specific KIF5B-RET transgenic mice by Cre recombinase-mediated cassette exchange (Cre-RMCE) to model the oncogenic activity of KIF5B- RET in lung type II alveolar cells in bitransgenic mice. In Specific Aim 1, plasmids carrying a tetO-KIF5B-RET cassette flanked by heterospecific L2/L3 loxP sites and Cre will be co-injected into fertilized eggs (zygotes) derived from different lines of existing Cre-RMCE-capable transgenic mice to replace the original floxed tetO- SHP2 transgenes with tetO-KIF5B-RET. In Specific Aim II, inducible KIF5B-RET expression and effects of KIF5B-RET on lung tumor development and maintenance will be evaluated in CCSP-rtTA/tetO-KIF5B-RET bitransgenic mice. Since our Cre-RMCE-capable SHP2 transgenic mice have already been characterized for inducible expression and function of the transgene in the lung, new transgenic mice derived from these lines by cassette exchange are expected to display reliable inducible transgene expression. This will accelerate the generation and examination of novel useful transgenic mouse models. Thus, the study will generate genetically engineered animals urgently needed for development of new therapy targeting the KIF5B-RET molecular subtype of lung adenocarcinoma. Furthermore, the study will identify the most efficient Cre-RMCE- capable transgenic line(s) harboring a single copy of the KIF5B-RET transgene for subsequent studies to derive new transgenic mice. This will provide an important resource for the research community to accelerate the generation of mouse models of newly identified molecular lesions.